SYSTEMATIC REVISION OF THE FISH BLOOD FLUKES WITH DIAGNOSES OF CHIMAEROHEMECIDAE YAMAGUTI, 1971, ACIPENSERICOLIDAE N. FAM., SANGUINICOLIDAE POCHE, 1926, ELOPICOLIDAE N. FAM., AND APOROCOTYLIDAE ODHNER, 1912

Abstract

We herein morphologically diagnose the 5 natural groups of fish blood flukes and name them. Species of Chimaerohemecidae Yamaguti, 1971 infect chimeras, sharks, and rays (Chondrichthyes) and have C-shaped lateral tegumental spines and a non-sinusoidal testis or lack spines and have a sinusoidal testis. Species of Acipensericolidae n. fam. infect sturgeons and paddlefish (Acipenseriformes) and have a robust, bowl-shaped, pedunculate anterior sucker, lateral tegumental spines that are spike-like (not C shaped), an inverse U-shaped intestine (anterior ceca absent) with posterior ceca terminating near the excretory bladder, 6 testes (inter-cecal ovoid or oblong, lacking deep lobes; including 1 post-ovarian testis), a Laurer’s canal, and a dextral common genital pore. Species of Sanguinicolidae Poche, 1926 infect primarily later-branching freshwater ray-finned fishes (Teleostei) and have a diminutive anterior sucker, a medial esophageal swelling (pouch), short, radial ceca of approximately equal length or short anterior ceca plus an elongate, dendritic posterior cecum, testis with appendix-like lateral lobes, no Laurer’s canal, and separate or common genital pores. Species of Elopicolidae n. fam. infect ladyfishes, tarpons, and catadromous eels (Elopomorpha) and have a robust, bowl-shaped, pedunculate anterior sucker, lateral tegumental spines that are spike-like (can be lost in adult), short or indistinct anterior ceca, posterior ceca that terminate at level of the testis(es), a single testis or 2 testes, a Laurer’s canal present or absent, and a sinistral common genital pore and atrium. Species of Aporocotylidae Odhner, 1912 primarily infect later-branching marine and estuarine ray-finned fishes (Teleostei) and have a spheroid anterior sucker with concentric rows of circumferential spines or the spheroid anterior sucker is lost in adults or adults have a diminutive anterior sucker, a sinuous esophagus lacking a pouch, an X- or H-shaped intestine having 4 ceca, long anterior ceca (or secondarily lost), smooth posterior ceca that extend posteriad in parallel with respective body margin and terminate near the posterior body end, testis(es) that lack appendix-like lateral lobes, no Laurer’s canal, and a sinistral common genital pore or separate genital pores that are sinistral. Our 28S phylogeny recovered the fish blood flukes as monophyletic and each of the morphologically diagnosed families as monophyletic and sister to the remaining blood flukes infecting turtles and homeotherms. Acipensericolidae was recovered sister to the clade comprising Chimaerohemecidae + Sanguinicolidae and Elopicolidae + Aporocotylidae. The branching order and interrelationships of these families remains unsettled perhaps because of low taxon sampling among non-aporocotylids and extinction of intermediate taxa.

The fish blood flukes, “Aporocotylidae Odhner, 1912” (see Bullard et al., 2009), presently comprise 5 morphologically distinct but as of yet undiagnosed lineages, 46 accepted genera, and >175 spp. Yamaguti (1958, 1971) proposed several subfamilies for the relatively few genera that were accepted at the time; however, none of those subfamilies were adopted because their diagnoses were based on erroneous character state assignments or homoplasy and because they are routinely and unequivocally recovered as paraphyletic or polyphyletic (Bullard et al., 2006, 2008, 2012; Orélis-Ribeiro et al., 2014; Orélis-Ribeiro and Bullard, 2015, 2016; Warren et al., 2017, 2019, 2020, 2021, 2023; Warren and Bullard, 2019, 2021). Since Smith (1997a, 1997b, 2002) reviewed the 20 genera of fish blood flukes that were accepted in 2002, an additional 26 genera and >100 species have been described. This later body of work collectively included many new taxonomic characters and character states with which to diagnose, differentiate, and classify fish blood fluke genera and the 5 main clades they comprise (Bullard and Overstreet, 2003, 2008; Bullard et al., 2006, 2008, 2012; Bullard and Jensen, 2008; Bullard, 2010a, 2010b, 2014; Orélis-Ribeiro et al., 2013, 2014, 2017; Truong and Bullard, 2013; Orélis-Ribeiro and Bullard, 2015, 2016; Cribb et al., 2017; Warren et al., 2017, 2019, 2020, 2021, 2023; Warren and Bullard, 2019, 2020; Cutmore et al., 2022). We herein diagnose these lineages using morphology, propose new families for them, and test this new classification scheme using a 28S phylogeny.

MATERIALS AND METHODS

The new and elevated family group names herein follow the recommendations of the International Commission on Zoological Nomenclature (ICZN, 1999; Notton et al., 2011). Specifically, Article 29.1, Formation of family-group names, states that “A family-group name is formed by adding to the stem of the name [Art. 29.3] of the type genus, or to the entire name of the type genus [see Article 29.6], a suffix as specified in Article 29.2” (Yamaguti, 1958, 1971). Article 29.3, Determination of stem in names of type genera, states that “The stem of a family-group name is based on the name of its type genus [Art. 63]…” Article 36, Principal of coordination, states that “A name established for a taxon at any rank in the family group (which includes the subfamily, family, and superfamily) is deemed to have been simultaneously established for nominal taxa at all other ranks in the family group; all these taxa have the same type genus, and their names are formed from the stem of the name of the type genus [Art. 29.3] with appropriate change of suffix [Art. 34.1]. The name has the same authorship and date at every rank.” Article 64, Choice of type genus, states that “An author who wishes to establish a new nominal family-group taxon may choose as type genus any included nominal genus the name of which he or she regards as valid [Art. 11.7.1], not necessarily that having the oldest name. The choice of type genus determines the stem of the name of the nominal family-group taxon [Art. 29.1].”

Each family diagnosis herein includes the available information on the known/reported intermediate and definitive hosts, site of infection in the definitive host, cercarial morphology, egg shape and site of infection, schistosomulum morphology, and adult morphology. For some of the proposed families, the life cycles of none, few, or several of the species are known.

Phylogenetic methods follow that of Warren and Bullard (2019), Bullard and Dutton (2022), and Whelan et al. (2022) (Table I). Members of Liolopidae Dollfus, 1934, Diplostomoidea Poirier, 1886, and Brachylaimoidea Joyeuz and Foley, 1930 were selected as functional outgroup taxa for the Schistosomatoidea Stiles and Hassal, 1898 (see De León and Hernández-Mena, 2019; Bullard and Dutton, 2022). All taxa included in the phylogenetic analysis have been published with morphological descriptions except the following unpublished sequences of ours: Sanguinicolidae sp. from heart of Micropterus sp. collected in the Coosa River, Alabama; Sanguinicolidae sp. from the body wash of Mystus cf. mysticetus in the Mekong River, Vietnam; Sanguinicolidae sp. from the body cavity of Pangasius cf. macronema collected in the Mekong River, Vietnam; Chimaerohemecidae sp. from the gill epithelium of the smooth butterfly ray, Gymnura micrura (Bloch and Schneider, 1801) collected in Mobile Bay, Alabama; Chimaerohemecidae sp. from the heart of smalltooth sawfish, Pristis pectinata Latham, 1796 collected in the Gulf of Mexico; and Chimaerohemecidae sp. from the heart of Gymnura cf. poecilura collected in the South China Sea off Vietnam. The specimens that produced the sequences we labeled herein as “Sanguinicolidae sp.” (several species; Fig. 1; Table I) had the diagnostic features for the family as diagnosed herein. Those of “Chimaerohemecidae sp.” were diagnosed by having C-shaped lateral tegumental spines (autopomorphic for that family) with the exception of that from the smooth butterfly ray. That sequence is by definition a nonugen (Roberts et al., 2018) and was sourced from blood fluke eggs in the gill. No family-level diagnostic features for blood fluke miracidia have been proposed, but we conservatively identified the eggs as a species of the family based on the sequence being recovered within other morphologically identified sequences of that family.

Table I.

New 28S sequences generated for the present study. Superscripts indicate specimens in Figure 1.

Species	Host	Locality	GenBank accession nos.
Chimaerohemecidae sp.13	Longtail butterfly ray, Gymnura cf. poecilura (Shaw, 1804) Dor, 1984 (Myliobatiformes: Gymnuridae)	Eastern Sea, off Nha Trang, Vietnam	OQ709101
Chimaerohemecidae sp.12	Smalltooth sawfish, Pristis pectinata Latham, 1796 (Rhinopristiformes: Pristidae)	Eastern Gulf of Mexico, off Naples, Florida	OQ709102
Chimaerohemecidae sp.11	Smooth butterfly ray, Gymnura micrura (Bloch and Schneider, 1801) Uyeno and Miyake, 1983 (Myliobatiformes: Gymnuridae)	Northern Gulf of Mexico, Mobile Bay, Alabama	OQ709103
Sanguinicolidae sp.4	Bass, Micropterus sp. (Lacepède 1802) Berg, 1949 (Centrarchiformes: Centrarchidae)	Coosa River, Alabama	OQ709104
Sanguinicolidae sp.6	Mystus cf. mysticetus Roberts, 1992 (Siluriformes: Bagridae)	Dong Thap fish market (Mekong River), Vietnam	OQ709105
Sanguinicolidae sp.7	Pangasius cf. macronema Bleeker, 1850 (Siluriformes: Pangasiidae)	Dong Thap fish market (Mekong River), Vietnam	OQ709106
Nomasanguinicola canthoensis Truong and Bullard, 2013	Broadhead catfish, Clarias macrocephalus Günther 1864 (Siluriformes: Clariidae)	Can Tho fish market, (Mekong River), Vietnam	OQ709107
Hyperandrotrema walterboegeri Orélis-Ribeiro and Bullard, 2013	Shortfin mako shark, Isurus oxyrinchus Rafinesque, 1810 (Lamniformes: Lamnidae)	Northern Gulf of Mexico, off Dauphin Island, Alabama	OQ709108
Myliobaticola richardheardi Bullard and Jensen, 2008	Atlantic stingray, Hypanus sabinus (Lesueur, 1824) Last, Manjaji-Matsumoto, Naylor, and White, 2016 (Myliobatiformes: Dasyatidae)	Mississippi Sound, northern Gulf of Mexico, off Biloxi, Mississippi, USA	OQ709109
Selachohemecus benzi Bullard, Overstreet, and Carlson, 2006	Blacktip shark, Carcharhinus limbatus (Valenciennes, 1839) Compagno, 1973 (Carcharhiniformes: Carcharhinidae)	Northern Gulf of Mexico, Mississippi,	OQ709110
Selachohemecus olsoni Short, 1954	Atlantic sharpnose shark, Rhizoprionodon terraenovae (Richardson, 1837) Springer, 1964 (Carcharhiniformes: Carcharhinidae)	Mississippi Sound, northern Gulf of Mexico, Mississippi,	OQ709111

Figure1.

Maximum-likelihood phylogeny based on the large subunit ribosomal DNA (28S). Labels in front of nodes are bootstrap support values. Values <80 not shown. New sequences generated along with GenBank accession numbers for this study in Table I. Scale bar indicates substitutions per site. Sequences for blood flukes not yet described or cercarial sequences follow associated numbering: ¹Sanguinicolidae spp., cercariae ex. Posticobia brazieri (Smith 1882), Brisbane River, Queensland, Australia (Cutmore et al., 2022); ²Sanguinicolidae sp. W5003, cercaria ex. Plotiopsis balonnensis (Conrad, 1850), Northern Territory, Victoria River, Australia (Brant et al., 2006); ³Sanguinicolidae sp. ex Juga plicifera (Lea, 1838), King, Hills, Dell, Taylor Creek; McKenzie River, Oregon (Preston et al., 2020); ⁴Sanguinicolidae sp. ex Micropterus sp. (Lacepède 1802), Coosa River, Alabama; ⁵Sanguinicolidae sp. W5004, cercaria ex. Amerianna carina (Adams, 1861), Northern Territory, Mary River Floodplain, Australia (Brant et al., 2006); ⁶Sanguinicolidae sp. ex. Mystus cf. mysticetus, Mekong River, Vietnam; ⁷Sanguinicolidae sp. ex. Pangasius cf. macronema, Mekong River, Vietnam; ⁸Chimaerohemecidae sp., cercaria ex. Plebidonax deltoides (Lamarck, 1818), Stockton Beach, New South Wales, Australia (Cribb et al., 2017); ⁹Chimaerohemecidae sp. H, cercaria ex. Megapitaria squalida (G. B. Sowerby I, 1835), off Santa Rosalía, Gulf of California, Mexico Gulf of California, Mexico (Cutmore et al., 2022); ¹⁰Chimaerohemecidae sp., cercaria ex. Solen viridis (Say, 1821), Gulf of Mexico (Warren and Bullard, 2019); ¹¹Chimaerohemecidae sp. ex. Gymnura micrura (Bloch and Schneider, 1801), Mobile Bay, Alabama; ¹²Chimaerohemecidae sp. ex. Pristis pectinata Latham, 1796, Gulf of Mexico; ¹³Chimaerohemecidae sp. ex. Gymnura cf. poecilura, Nha Trang, Vietnam; ¹⁴Schistosomatoidea sp. W1134 ex. Biomphalaria sudanica (E von Martens, 1870), Queen Elizabeth National Park, Uganda (Brant et al., 2006); ¹⁵Schistosomatoidea sp. W1284 ex. Segmentorbis kanisaensis (Preston, 1914), Lake Victoria, Kenya (Brant et al., 2006).

DESCRIPTIONS

Chimaerohemecidae Yamaguti, 1971

Accepted genera:

Chimaerohemecus Van der Land, 1967 [type]; Selachohemecus Short, 1954; Orchispirium Madhavi and Rao, 1970; Hyperandrotrema Maillard and Ktari, 1978; Myliobaticola Bullard and Jensen, 2008; Ogawaia Cutmore, Cribb, and Yong, 2018; Gymnurahemecus Warren, Ruiz, Whelan, Kritsky, and Bullard, 2019; Electrovermis Warren and Bullard, 2019; Achorovermis Warren and Bullard, 2020; Aetohemecus Warren and Bullard, 2021; Homestios Warren and Bullard, 2021.

Diagnosis:

Hermaphroditic, asexual reproduction in marine bivalves (Electrovermis), lacking encysted metacercaria or second intermediate host (Table II). Sporocyst spheroid or ovoid, with few cercariae. Cercaria non-acetabulate apharyngeate, non-ocellate; spinous anterior sucker present; tegumental body spines distributing as lateral transverse rows; body fin present; tail brevifurcate; furcae asymmetrical, lacking fins; discernable gonads or genitalia indistinct. Extrauterine eggs embedding within and undergoing considerable larval development in gill epithelium of definitive host.

Table II.

Reports of intermediate hosts and completed life cycles of fish blood flukes.

Host	Cercaria	Locality	Reference
Gastropoda
Amerianna carinata (H. Adams, 1861) (Hygrophila: Planorbidae); Glyptophysa gibbose (A. Gould, 1847) (Hygrophila: Planorbidae)	Sanguinicolidae sp.5 (W5004)	Northern Territory, Mary River Floodplain, Australia	Brant et al. (2006)
Biomphalaria sudanica (E von Martens, 1870) (Hygrophila: Planorbidae)	Schistosomatoidea sp.14 (W1134)	Queen Elizabeth National Park, Uganda	Brant et al. (2006)
Campeloma decisum (Say, 1817) (Architaenioglossa: Viviparidae)	Pseudosanguinicola occidentalis	New York	Bacha (1966)
Fluminicola seminalis (Hinds, 1842) (Littorinimorpha: Hydrobiidae)	Sanguinicola klamathensis	Darrah Springs Hatchery, Tehama Co., California	Wales (1958)
Fluminicola virens (I.Lea, 1838) (Littorinimorpha: Hydrobiidae)	Sanguinicola idahoensis	Clearwater River, Clearwater Co., Idaho	Schell (1974)
Heleobia australis (Orbigny, 1835) (Littorinimorpha Cochliopidae)	Aporocotylidae gen sp. 1	Arroyo Cangrejo, Buenos Aires province, Argentina	Matías et al. (2014)
Juga plicifera (Lea, 1838) (as Oxytrema silicula) (Caenogastropoda: Semisulcospiridae)	Sanguinicola alseae	Alsea River, Benton County, Oregon	Meade and Pratt (1965)
Juga plicifera (Lea, 1838) (as Oxytrema silicula) (Caenogastropoda: Semisulcospiridae)	Sanguinicolidae sp.3 (Aporocotylidae sp.)	King, Hills, Dell, Taylor Creek; McKenzie River	Preston et al. (2020)
Ancylus fluviatilis O. F. Müller, 1774 (Hygrophila: Planorbidae)	Sanguinicola rutili	Salamanca, Spain	Simon-Martin et al. (1987)
Leptoxis carinata (Bruguiere, 1792) “Oxytrema circumlineata” (Caenogastropoda: Pleuroceridae)	Sanguinicola fontinalis	Susquehanna River, Harrisburg, Pennsylvania	Hoffman et al. (1985)
	Sanguinicola davisi	Darrah Springs Hatchery, Tehama Co., California	Wales (1958)
	Sanguinicola klamathensis	Klamath River, Oregon	Meade (1967)
Littorodinops monroensis Frauenfeld, 1863 (Littorinimorpha: Hydrobiidae)	Aporocotylidae sp. cercaria type 1	Northern Gulf of Mexico, Grand Lagoon, Florida	Bullard (2007)
Lymnaea stagnalis (Linnaeus, 1758) (Hygrophila: Lymnaeidae)	Sanguinicola sp.	Lake Glubokoye, Russia	Nikolaeva (1985)
Peregriana peregra (O. F. Müller, 1774) (as Lymnaea) (Hygrophila: Lymnaeidae)	Sanguinicola inermis	Europe	Kirk and Lewis (1993)
Planorbella trivolvis Say, 1817) (as Helisoma) (Hygorphila: Planorbidae)	Cercaria brevifurca	Vicinity of St. Louis, Missouri	McCoy (1929)
	Cercaria whitentoni	Stillwater Tourist Park, Stillwater, Oklahoma	Croft (1933)
Plotiopsis balonnensis (Conrad, 1850) (Caenogastropoda: Thiaridae) (as Thiara balannensis)	Sanguinicolidae sp.2 (W5003)	Northern Territory, Victoria River, Australia	Brant et al. (2006)
Posticobia brazieri (Smith 1882) (Littorinimorpha: Tateidae)	Paracardicoloides yamagutii	Downfall Creek, Wivenhoe Pocket, Queensland, Australia	Nolan and Cribb (2004)
	Sanguinicolidae sp.1 (Aporocotylidae sp. B)	Unnamed tributary of Brisbane River, Australia; Fairnie Brook, Queensland, Australia	Cutmore et al. (2022)
	Sanguinicolidae sp.1 (Aporocotylidae sp. C)	Moggill Creek, Queensland, Australia; unnamed tributary of Brisbane River, Queensland, Australia	Cutmore et al. (2022)
	Sanguinicolidae sp.1 (Aporocotylidae sp. D)	Unnamed tributary of Brisbane River, Queensland, Australia	Cutmore et al. (2022)
	Sanguinicolidae sp.1 (Aporocotylidae sp. E)	Unnamed tributary of Brisbane River, Queensland, Australia	Cutmore et al. (2022)
	Sanguinicolidae sp.1 (Aporocotylidae sp. F)	Moggill Creek, Queensland, Australia	Cutmore et al. (2022)
	Sanguinicolidae sp.1 (Aporocotylidae sp. G)	Churchbank Weir, Queensland, Australia	Cutmore et al. (2022)
Segmentorbis kanisaensis (Preston, 1914) (Hygrophila: Planorbidae)	Schistosomatoidea sp.15 (W1284)	Lake Victoria, Kenya	Brant et al. (2006)
Valvata tricarinata (Say, 1817) (Heterobranchia: Valvatidae)	Sanguinicola lophophora	Lake Francis, Isanti Co., Minnesota	Erickson and Wallace (1959)
Bivalvia
Anadara trapezia (Deshayes, 1839) (Arcida: Arcidae)	Elopicola bristowi	Eastern Moreton Bay, Queensland, Australia	Cutmore et al. (2022)
Argopecten irradians (Lamarck, 1819) (Pectinida: Pectinidae)	Cercaria martini	Northwestern Atlantic Ocean, Woods Hole, Massachusetts	Linton (1915); Stunkard (1983)
Chione cancellata (Linnaeus, 1767) (Venerida: Veneridae)	Cercaria cristulata	Northern Gulf of Mexico, Alligator Point, Florida	Holliman (1961)
Donax variabilis Say, 1822 (Cardiida: Donacidae)	Cercaria asymmetrica	Northern Gulf of Mexico, Alligator Point, Florida	Holliman (1961)
	Electrovermis zappum	Northern Gulf of Mexico, Fort Morgan, Alabama	Warren and Bullard (2019)
Ensis macha (Molina, 1782) (Adapedonta: Pharidae)	Aporocotylidae sp. cercaria	La Tapera, San Matias Gulf, Patagonia, Argentina	Vázquez et al. (2013); Orellana and Lohrmann (2015)
Eucallista purpurata (Lamarck, 1818) (Venerida, Veneridae) (as Amiantis purpurata)	Aporocotylidae sp. cercaria	El Molino Beach, San Matias Gulf, Patagonia, Argentina	Gilardoni et al. (2011); Carvalho et al. (2015)
Mactra isabelleana d’Orbigny, 1846 (Venerida: Mactridae)	Aporocotylidae sp. cercaria	Cassino Beach, San Matias Gulf, Patagonia, Argentina	Carvalho et al. (2015)
Megapitaria squalida (G. B. Sowerby I, 1835) (Venerida, Veneridae)	Chimaerohemecidae sp.9 (Aporocotylidae sp. H)	Off Santa Rosalía, Gulf of California, Mexico	Yee-Duarte et al. (2017); Cutmore et al. (2022)
Mercenaria capechiensis (Gmelin, 1791) (Venerida: Veneridae)	Cercaria mercenariae	Western Gulf of Mexico, Galveston Island, Texas	Wardle (1979)
Mesodesma donacium (Lamarck, 1818) (Venerida: Mesodesmatidae)	Aporocotylidae sp. cercaria	Carelmapu, Chile	López et al. (2014)
Plebidonax deltoides (Lamarck, 1818) (as Donax) (Cardiida: Psammobiidae)	Chimaerohemecidae sp.8 (Aporocotylidae sp. NSW1)	Stockton Beach, New South Wales, Australia	Cribb et al. (2017)
Solemya velum Say, 1822 (Solemyida: Solemyidae)	Cercaria solemyae	Northwestern Atlantic Ocean, Woods Hole, Massachusetts	Martin (1944)
Solen viridis (Say, 1821) (Adapedonta: Solenidae)	Chimaerohemecidae sp.10 (Aporocotylidae sp. cercaria type 2)	Northern Gulf of Mexico, Mississippi Sound, Mississippi	Bullard (2007); Warren and Bullard (2019)
Tagelus divisus (Spengler, 1794) (Cardiida: Solecurtidae)	Aporocotylidae sp. cercaria	Northwestern Atlantic Ocean, Biscayne Bay, Florida	Fraser (1967)
Annelida
Amphicteis gunneri (Sars, 1835) (Terebellida: Ampharetidae)	Cercaria amphicteis	Northern Gulf of Mexico, Apalachicola River, Florida	Oglesby (1961)
Amphitrite ornata (Leidy, 1855) (Terebellida: Terebellidae)	Cardicola parvus	Oyster landing in North Inlet, near Georgetown, South Carolina and Charleston, South Carolina	Siegel et al. (2018)
Amphitrite sp. Müller, 1771 (Terebellida: Terebellidae)	Cardicola forsteri	Kushimoto, Wakayama Prefecture, Japan	Shirakashi et al. (2016)
Artacama proboscidea Malmgren, 1866 (Terebellida: Terebellidae)	Aporocotyle simplex	Øresund, north of the island Veen, between Denmark and Sweden	Køie (1982)
Enoplobranchus sanguineus (Verrill, 1873) (Terebellida: Terebellidae)	Cardicola parvus	Oyster landing in North Inlet, near Georgetown, South Carolina and Charleston, South Carolina	Siegel et al. (2018)
Hydroides dianthus Verrill, 1873 (Sabellida: Serpulidae)	Cercaria loossi	Northwestern Atlantic Ocean, Woods Hole, Massachusetts	Linton (1915)
Lanassa nordenskioldi Malmgren, 1866 (Terebellida: Terebellidae)	“…morphologically indistinguishable” from Aporocotyle simplex	Seyðisfjörður, Eastern Iceland	Køie and Peterson (1988)
Lanicides vayssierei (Gravier, 1911) (Terebellidae: Terebellidae)	Cercaria hartmanae	Northwestern Atlantic Ocean, Woods Hole, Massachusetts	Martin (1952)
Longicarpus modestus (Quatrefages, 1866) (Terebellida: Terebellidae)	Cardicola forsteri	Off Port Lincoln, South Australia	Cribb et al. (2011)
Nicolea gracilibranchis (Grube, 1878) (Terebellida: Terebellidae)	Cardicola orientalis	Kushimoto, Wakayama Prefecture, Japan	Shirakashi et al. (2016)
Terebella lapidaria Linnaeus, 1767 (Terebellida: Terebellidae)	Cardicola laruei	Oyster landing in North Inlet, near Georgetown, South Carolina and Charleston, South Carolina	Siegel et al. (2018)
Terebella sp. Linnaeus, 1767 (Terebellida: Terebellidae)	Cardicola opisthorchis	Off Tsushima, Nagasaki, Japan	Sugihara et al. (2014)

Body of adult dorsoventrally flat, ovoid or elongate, spinose and having each spine mounted on a muscular peduncle (Aetohemecus, Chimaerohemecus, Gymnurahemecus, Hyperandrotrema, Selachohemecus) or aspinose and lacking peduncles or tubercles (Achorovermis, Electrovermis, Homestios, Myliobaticola, Ogawaia) or having aspinose lateral tubercles (Orchispirium); spine bosses, posterolateral body protuberance, head collar, copulatory bursa, acetabulum, and median esophageal pouch or plicate organ lacking. Tegumental spines (if present) robust, C shaped, each mounted on a muscular peduncle, directed ventrally, distributing in a single lateral column (Aetohemecus, Gymnurahemecus, Selachohemecus) or 2 lateral columns (Chimaerohemecus) or a lateral field of interspaced spines (Hyperandrotrema). Rosethorn-shaped spines absent. Sensory papillae absent. Anterior sucker nearly indistinct, aspinose, diminutive, not pedunculate, not bowl shaped; mouth subterminal, a minute pore-like opening. Nervous system obvious depending on species; dorsolateral and ventrolateral nerve cords extending for nearly entire body length, forming anterior and posterior nerve commissures; secondary lateral branches obvious or indistinct. Pharynx absent. Esophagus long, sinuous or straight, medial, having a medial swelling (esophageal bulb) and/or posterior swelling (chamber) immediately anterior to cecal bifurcation; esophageal gland surrounding esophagus for entire length or limited to middle or posterior portion of esophagus (indistinct in Aetohemecus, Homestios, Myliobaticola). Intestine inverse U shaped or (secondarily) X shaped (Selachohemecus, Aetohemecus), bifurcating at midline, not crossing opposing cecum; posterior ceca slightly asymmetrical (Aetohemecus, Chimaerohemecus, Gymnurahemecus, Homestios, Hyperandrotrema, Myliobaticola, Orchispirium, Selachohemecus) or markedly asymmetrical (Achorovermis, Electrovermis, Ogawaia), blind ended, paired, terminating in anterior half of body (Achorovermis, Aetohemecus, Electrovermis, Gymnurahemecus, Homestios, Myliobaticola, Selachohemecus, Ogawaia) or posterior body extremity (Chimaerohemecus, Hyperandrotrema, Orchispirium). Genitalia occupying posterior one-third of body, post-gonadal.

Testis single, having smooth borders or exceptionally having posteriorly directed lobes (Orchispirium), non-sinusoidal (Aetohemecus, Chimaerohemecus, Gymnurahemecus, Hyperandrotrema, Selachohemecus) or sinusoidal (Achorovermis, Electrovermis, Homestios, Myliobaticola, Ogawaia, Orchispirium), anterior to ovary and genitalia, post-cecal (Achorovermis, Aetohemecus, Electrovermis, Gymnurahemecus, Homestios, Myliobaticola, Ogawaia, Selachohemecus) or inter-cecal (Chimaerohemecus, Hyperandrotrema, Orchispirium), not extending anterior to cecal bifurcation. Vasa efferentia coalescing ventrally and in posterior portion of testis, uniting to form vas deferens in posterior margin of testis; vas deferens straight (Achorovermis, Chimaerohemecus, Hyperandrotrema, Selachohemecus, Orchispirium, Ogawaia) or curved (Aetohemecus, Gymnurahemecus, Homestios) or looped (Electrovermis). Cirrus sac present, enveloping internal seminal vesicle and cirrus, post-gonadal or lateral to ovary (Aetohemecus, Orchispirium). Auxiliary external seminal vesicle absent.

Female reproductive system comprising a single ovary, oviduct, seminal receptacle that can comprise part of oviduct (oviducal seminal receptacle) (Myliobaticola, Orchispirium, Selachohemecus) or oviducal ampullae (Aetohemecus, Gymnurahemecus, Hyperandrotrema), vitellarium and primary vitelline collecting duct, oötype, uterus, and metraterm. Ovary single, wholly inter-cecal (Chimaerohemecus, Hyperandrotrema) or wholly post-cecal (Achorovermis, Aetohemecus, Electrovermis, Gymnurahemecus, Homestios, Myliobaticola, Ogawaia, Orchispirium, Selachohemecus), primarily post-testicular medial, or lateral to midline (Gymnurahemecus, Orchispirium, Selachohemecus), deeply lobed (Aetohemecus, Chimaerohemecus, Gymnurahemecus, Homestios, Hyperandrotrema, Selachohemecus) or superficially lobed (Electrovermis) or a loose aggregation of ova (Achorovermis, Myliobaticola, Orchispirium). Oviduct a thin-walled duct emanating from ovary, straight to sinuous (Aetohemecus, Chimaerohemecus, Gymnurahemecus, Selachohemecus, Ogawaia, Orchispirium) or looping (Achorovermis, Myliobaticola); Hyperandrotrema walterboegeri has a straight to sinuous oviduct and H. cetorhini has a looping oviduct. Indistinct in Electrovermis and Homestios. Vitellarium follicular, diffuse (Achorovermis, Electrovermis, Myliobaticola), in dense lobules (Aetohemecus, Gymnurahemecus, Homestios), or granular (Selachohemecus, Ogawaia, Orchispirium), symmetrical (Aetohemecus, Electrovermis, Myliobaticola, Ogawaia) or asymmetrical (Achorovermis, Chimaerohemecus, Gymnurahemecus, Homestios, Orchispirium, Selachohemecus; H. walterboegeri is asymmetrical and H. cetorhini is symmetrical), filling space from nerve commissure to proximal portion of testis (Achorovermis, Homestios), ovary (Aetohemecus, Gymnurahemecus, Selachohemecus), or genital pore (Chimaerohemecus, Electrovermis, Hyperandrotrema, Myliobaticola, Ogawaia, Orchispirium). Primary vitelline duct extending posteriad along dextral body margin or along midline, ventral to ovary (Chimaerohemecus), not forming a transverse vitelline duct or reservoir, uniting with oviduct in posterior body end proximal to and near oötype. Indistinct in Electrovermis, Homestios, and Myliobaticola. Laurer’s canal present (Chimaerohemecus, Hyperandrotrema, Gymnurahemecus) or absent (Aetohemecus, Achorovermis, Electrovermis, Homestios, Myliobaticola, Ogawaia, Orchispirium, Selachohemecus). Oötype post-cecal or exceptionally inter-cecal in Hyperandrotrema, posterior to common genital pore or exceptionally dextral to common genital pore (Orchispirium) or anterior to common genital pore (Selachohemecus), comprising an inconspicuous ovoid chamber; Mehlis’ gland indistinct, staining poorly and difficult to visualize with light microscopy of fixed, stained specimens. Uterus post-testicular or overlapping posterior margin of testis (Ogawaia), post-cecal or inter-cecal (Chimaerohemecus, Hyperandrotrema), typically straight or exceptionally convoluted (Aetohemecus, Chimaerohemecus, Hyperandrotrema, Ogawaia, Orchispirium), having ascending and descending portions; ascending portion extending anteriad along midline from oötype (Aetohemecus, Chimaerohemecus, Hyperandrotrema, Gymnurahemecus) or extending posteriad before turning anteriad and extending along midline (Achorovermis, Ogawaia, Selachohemecus), typically curving posteriad at level of testis (Aetohemecus, Ogawaia, Orchispirium), ovary (Achorovermis, Chimaerohemecus, Gymnurahemecus, Homestios, Hyperandrotrema, Myliobaticola, Selachohemecus), or seminal vesicle (Achorovermis); descending portion extending posteriad along sinistral body margin, uterus + metraterm flanking cirrus sac, metraterm muscularized to varying degrees or indistinct; uterine eggs thin-shelled, oblong (Aetohemecus, Achorovermis, Electrovermis, Homestios, Myliobaticola, Ogawaia, Orchispirium) or circular (Gymnurahemecus, Hyperandrotrema, Selachohemecus—S.A.B. observations), lacking filaments, lacking discernible miracidium, undergoing considerable larval development in tissue of definitive host (at least 1 species has eggs that hatch while in the gill of the fish host). Common genital pore dorsal, on sinistral body margin, post-gonadal aspinose, post-cecal or exceptionally inter-cecal in Hyperandrotrema; common genital atrium present in Chimaerohemecus and Hyperandrotrema, indeterminate for other genera.

Excretory vesicle minute, medial, having excretory arms or exceptionally lacking excretory arms (Gymnurahemecus) (indeterminate for Aetohemecus, Electrovermis, Myliobaticola, Ogawaia). Maturing in blood of sharks, rays, and chimaeras (Chondrichthyes).

Differential diagnosis:

Adults having C-shaped lateral tegumental spines each mounted on muscular peduncle and having non-sinusoidal testis or lacking lateral tegumental spines and having sinusoidal testis. Maturing in blood of sharks, rays, and chimaeras (Chondrichthyes).

Remarks

Based on morphology, we diagnose 2 groups within Chimaerohemecidae. One group matures in chimaeras, sharks, and epipelagic rays (Gymnuridae, Myliobatidae) and differs from all other blood flukes by having C-shaped lateral tegumental spines and a non-sinusoidal testis (Fig. 1). The other chimaerohemecids infect benthic rays (Narcinidae, Prisitidae, Glaucostegidae, Dasyatidae) and differ from all other blood flukes by lacking spines and having a sinusoidal testis. Warren and Bullard (2021) theorized that chimaerohemecids could eventually be split into different groups (subfamilies) based on the presence or absence of lateral tegumental spines and testis shape. Noteworthy also is that the X-shaped intestine of Selachohemecus and Aetohemecus is homoplasy, not homologous to the X-shaped intestine of Sanguinicola spp., for example, evidently evolved from the inverse U-shaped intestine of other chimaerohemecids (Warren and Bullard, 2021; present study; Fig. 1).

Only 1 study has matched cercariae and adults of a chimaerohemecid in the bivalve and chondrichthyan hosts (Warren and Bullard, 2019): Electrovermis zappum Warren and Bullard, 2019 infects the variable coquina clam, Donax variabilis Say, 1812 (Donacidae Fleming, 1828) and matures in the lesser electric ray, Narcine bancroftii (Griffith and Smith, 1834) (Narcinidae) in high-energy open-beach habitat in the northern Gulf of Mexico. Other studies have collected trematode cercariae from bivalves, sequenced them, conducted a phylogenetic analysis, inferred their taxonomic identity from the resulting tree topology, and presumed the identity of the definitive host group based on their clade assignment; which assumes no possibility of host switching (Cribb et al., 2017; Cutmore et al., 2022). Although we do not know the definitive host for any chimaerohemecid cercarial sequence other than E. zappum, sequences of these innominate cercariae clade with E. zappum and other batoid chimaerohemecids that lack C-shaped spines and have a sinusoidal testis. The cercaria for a blood fluke in which adults have C-shaped spines and lack a sinusoidal testis has not been reported.

Yamaguti (1971) proposed Chimaerohemecinae Yamaguti, 1971 for Chimaerohemecus trondheimensis Van der Land, 1967 (see Van der Land, 1967; Yamaguti, 1971), but his subfamily diagnosis was erroneous, including objective errors regarding the absence of lateral tegumental spines and the structure of the testis (Orélis-Ribeiro et al., 2013; Van der Land, 1967). Herein, we elevated the subfamily name to family as per the recommendations of the ICZN despite the core problems of Yamaguti’s subfamily diagnosis.

Acipensericolidae n. fam.

Accepted genera:

Acipensericola Bullard, Snyder, Jensen, and Overstreet, 2008.

ZooBank registration:

urn:lsid:zoobank.org:act:3647841C-A71D-48C5-8534-DC28C48FA73A.

Etymology:

Acipensericolidae refers to the definitive host group.

Diagnosis:

No life cycle known. Body of adult dorsoventrally flat, strongly ventrally concave, spinose; posterolateral body protuberance, head collar, copulatory bursa, acetabulum, and median esophageal pouch or plicate organ lacking. Tegumental body spines distributing as lateral transverse rows, spike-like lacking recurved tip. Rosethorn-shaped spines absent. Sensory papillae abundant, occupying ventrolateral body surface between lateral nerve cord and body margin.

Anterior sucker bowl shaped, centered on mouth, demarcated from the anterior body end by peduncle, having minute spines on inner anteroventral surface only; mouth subterminal, a relatively large opening (cf. chimaerohemecids). Nervous system well developed; dorso- and ventrolateral nerve cords present, extending for nearly entire body length, meeting to form anterior and posterior nerve commissures; secondary lateral branches obvious or indistinct. Pharynx between anterior sucker and nerve commissure, highly muscular, intensely basophilic. Esophagus medial, extending straight before connecting with intestine. Intestine inverse U-shaped, bifurcating at midline, with long posterior ceca only and no anterior cecum, lacking secondary rami, not crossing opposing cecum, extending posteriad to near body end.

Testes inter-cecal 6 in number, comprising 5 pre-ovarian testes and 1 post-ovarian testis. Vasa efferentia entwining throughout testicular tissue, with anterior and posterior trunks linking testicular column and posterior-most testis; vas deferens straight. Cirrus sac dextral, post-ovarian enveloping seminal vesicle and cirrus; cirrus everting dorsally near dextral body margin. Common genital atrium present; common genital pore dorsal, opening on dextral body margin, inter-cecal or dorsal to dextral cecum, post-ovarian aspinose. Auxiliary external seminal vesicle absent.

Ovary single, medial, inter-cecal separating anterior column of testes from single posterior testis, immediately posterior to testicular column, deeply lobed, located within posterior one-fourth of body. Oviduct a thin-walled duct emanating from ovary, sinistral, functioning as oviducal seminal receptacle, extending to level of posterior testis before recurving before meeting oötype. Vitellarium diffuse, extending dorsal and ventral to gonads and ceca, not extending laterally beyond ventrolateral nerve cords; primary vitelline duct extending posteriad, sinistral, single/unpaired, not forming a transverse vitelline duct or reservoir, arcing sinistral to posterior testis in column, uniting with oviduct in posterior body end proximal to and near oötype. Laurer’s canal present. Oötype spherical, inter-cecal inter-testicular, ventral to ovary, medial to cirrus sac. Uterus long, ventral to ovary, inter-cecal having ascending and descending portions, convoluted, extending anterior from oötype to level of posterior portion of testicular column before curving posteriad, typically containing several large eggs; uterine eggs oblong, one-third uterus maximum width, lacking filaments, lacking mature miracidium. Genitalia occupying posterior one-fourth of body, inter-testicular lateral to ovary and oötype.

Excretory vesicle large, Y shaped, cradling posterior testis, thin walled; excretory pore dorsal, subterminal. Maturing in blood of sturgeons and paddlefish (Chondrostei: Acipenseriformes).

Differential diagnosis:

Anterior sucker robust, bowl shaped, pedunculate, having minute spines on inner anteroventral surface only. Lateral tegumental spines spike-like (not C shaped, lacking recurved tip); distributing in ventrolateral transverse rows, with distal end having a sharp tip protruding slightly from tegument. Pharynx present. Intestine inverse U shaped; anterior ceca absent; posterior ceca terminating near excretory bladder. Testes 6 in number (5 pre-ovarian testes plus 1 post-ovarian testis), inter-cecal having shallow lobes, non-sinusoidal Laurer’s canal present. Vitellarium symmetrical; primary vitelline collecting duct sinistral. Common genital pore dextral. Maturing in blood of sturgeons and paddlefishes (Acipenseriformes)

Remarks

Acipensericolids mature in sturgeons and paddlefish (Acipenseriformes) and are readily differentiated from all other fish blood flukes by the combination of having a robust, bowl-shaped, pedunculate anterior sucker, lateral tegumental spines that are spike-like (not C shaped), an inverse U-shaped intestine (anterior ceca absent) with posterior ceca terminating near the excretory bladder, 6 testes (inter-cecal ovoid or oblong, lacking deep lobes; including 1 post-ovarian testis), a Laurer’s canal, and a dextral common genital pore.

Elopicolidae n. fam.

Accepted genera:

Elopicola Bullard, 2014 (type genus) and Paracardicoloides Martin, 1974.

ZooBank registration:

urn:lsid:zoobank.org:act:79EFE8E4-FB25-4297-AA93-8FA90CEEEC01.

Etymology:

Elopicolidae refers to the definitive host group.

Diagnosis:

Hermaphroditic, asexual reproduction in freshwater gastropods (Paracardicoloides) and marine bivalves (Elopicola), lacking encysted metacercaria or second intermediate host. Sporocyst oblong, harboring few cercariae. Cercaria non-acetabulate apharyngeate, non-ocellate having spinous anterior sucker, tegumental body spines, body fin, and symmetrical furcae with fins, lacking discernable gonads or genitalia, undergoing considerable larval development in vertebrate definitive host. Extrauterine eggs embedding within and undergoing considerable larval development in gill epithelium of definitive host.

Body of adult dorsoventrally flat, thick-bodied, spinose (Paracardicoloides spp. and juveniles of Elopicola spp.) or aspinose (adults of Elopicola spp.), lacking posterolateral body protuberance, head collar, copulatory bursa, acetabulum, and median esophageal pouch or plicate organ lacking. Tegumental body spines (if present) straight, spike-like lacking recurved tip, not distributing as ventrolateral transverse rows, with tips enveloped by tegument. Rosethorn-shaped spines absent. Sensory papillae abundant, occupying ventrolateral body surface between lateral nerve cord and body margin. Anterior sucker bowl shaped, centered on mouth, demarcated from anterior body end by peduncle, aspinous; mouth subterminal, a relatively large opening (cf. chimaerohemecids). Nervous system well developed; dorso- and ventrolateral nerve cords present, extending for nearly entire body length, meeting to form anterior and posterior nerve commissures; secondary lateral branches indistinct. Sensory papillae present (Elopicola) or absent (Paracardicoloides), bearing cilia (Elopicola) or lacking cilia (Paracardicoloides), not distributing in regular dorsolateral bands. Pharynx present (Elopicola) or absent (Paracardicoloides), between anterior sucker and nerve commissure, muscular, intensely basophilic. Esophagus medial, extending straight before connecting with intestine. Intestine typically inverse U-shaped or with abbreviated lobes of cecum directing anteriad, bifurcating at midline, with long posterior ceca, terminating in middle one-third of body.

Testes single (Elopicola) or 2 in number (Paracardicoloides), inter-cecal (Elopicola) or post-cecal (Paracardicoloides), medial, deeply lobed, in posterior half of body. Vasa efferentia entwining throughout testicular tissue, coalescing in anterosinistral region of testis to form vas deferens (Elopicola) or comprising anterior and posterior trunks linking anterior and posterior testes (Paracardicoloides); vas deferens straight, not convoluted, meeting with cirrus sac. Cirrus sac sinistral, pre-ovarian enveloping internal seminal vesicle and cirrus; cirrus everting dorsally in sinistral body half (Paracardicoloides) or everting dorsally near midline (Elopicola), post-cecal. Common genital atrium present; common genital pore dorsal, near sinistral body margin, post-cecal pre-ovarian. Auxiliary external seminal vesicle absent.

Ovary medial, post-cecal inter-testicular (Paracardicoloides) or post-testicular (Elopicola), deeply lobed. Oviduct a thin-walled duct emanating from ovary, medial, post-cecal post-genital pore; oviducal seminal receptacle present. Vitellarium follicular, an extensive network of narrow interconnecting branching bands having granular vitelline material, extending laterad beyond ventrolateral nerve cords, occupying space from anterior nerve commissure posteriad to level of distal tips of posterior ceca; primary vitelline duct single/unpaired, not forming a transverse vitelline duct or reservoir, arcing along posterodextral margin of testis/anterior testis, connecting with oötype posteriorly (Paracardicoloides) or laterally (Elopicola). Laurer’s canal present (Paracardicoloides) or absent (Elopicola). Oötype oblong, pre-ovarian anterior to common genital pore (Paracardicoloides) or posterior to common genital pore (Elopicola). Uterus comprising short ascending and descending portions, not convoluted, pre-ovarian inter-cecal (Paracardicoloides) or post-cecal (Elopicola); uterine eggs ovoid; metraterm pre-ovarian medial to cirrus sac.

Excretory vesicle large, Y shaped; excretory pore dorsal, subterminal. Maturing in blood of elopomorphs (Teleostei: Elopomorpha).

Differential diagnosis:

Anterior sucker robust, bowl shaped, aspinose in adults, pedunculate. Lateral tegumental spines spikelike not C shaped, not distributing in ventrolateral transverse rows, with distal end having a sharp tip enveloped by tegument (adults of Elopicola spp. are aspinose); mouth comprising a large opening. Pharynx present (Elopicola) or absent (Paracardicoloides). Intestine generally inverse U-shaped; anterior ceca short or nearly indistinct; posterior ceca terminating at level of testis (Elopicola)/anterior-most testis (Paracardicoloides). Testis(es) deeply lobed, non-sinusoidal comprising 1 pre-ovarian and inter-cecal testis plus 1 post-ovarian and post-cecal testis (Paracardicoloides) or comprising 1 inter-cecal testis (Elopicola); common genital atrium and pore present, sinistral, pre-ovarian. Vitellarium symmetrical; primary vitelline collecting duct dextral. Laurer’s canal present (Paracardicoloides) or absent (Elopicola). Egg in gill epithelium of fish host having elongate polar filaments. Maturing in blood of elopomorphs (Teleostei: Elopomorpha).

Remarks

Elopicolids mature in fishes having a leptocephalus larva: ladyfishes (Elops spp.), tarpons (Megalops spp.), and catadromous eels (Anguilla spp.) (all Elopomorpha). They differ from all other blood flukes by the combination of having a robust, bowl shaped, pedunculate anterior sucker, lateral tegumental spines that are spike-like (can be lost in adult), short or nearly indistinct anterior ceca, posterior ceca that terminate at level of the testis(es), a single testis or 2 testes, a Laurer’s canal present or absent, and a sinistral common genital pore and atrium.

Sanguinicolidae Poche, 1926

Accepted genera:

Sanguinicola Plehn, 1905 (type); Plehniella Szidat, 1951; Nomasanguinicola Truong and Bullard, 2013; Kritsky Orélis-Ribeiro and Bullard 2016; Pseudosanguinicola Warren and Bullard, 2023; and Cladocaecum Orélis-Ribeiro and Bullard 2016; we provisionally include Parasanguinicola Herbert and Shaharom-Harrison, 1995.

Diagnosis:

Hermaphroditic, asexual reproduction in freshwater gastropods (Sanguinicola), lacking encysted metacercaria or second intermediate host, having migratory larva (schistosomulum) infecting lymphatic and blood vascular system, having adult infecting blood and body cavity of vertebrate definitive host. Sporocyst present (Sanguinicola) or sporocyst and redia present (Sanguinicola). Sporocyst spheroid (Sanguinicola) or ovoid (Sanguinicola), having few or having many cercariae (Sanguinicola). Cercaria non-acetabulate apharyngeate, non-ocellate having spinous anterior sucker; tegumental body spines present or absent; body fin present or absent; lacking discernable gonads or genitalia; tail furcae symmetrical, fins present or absent; undergoing considerable larval development in vertebrate definitive host; extrauterine eggs embedding in fish gill epithelium and undergoing considerable larval development; miracidium can hatch and emerge from eggs embedded in fish gill epithelium.

Body of adult dorsoventrally flat, ventrally concave, aspinose or spinose (Sanguinicola, Parasanguinicola, Pseudosanguinicola); head collar, copulatory bursa, acetabulum, and median esophageal pouch or plicate organ lacking. Tegumental body spines straight, elongate, not distally recurved, deeply rooted in tegument (Sanguinicola) or only slightly protruding from tegument (Sanguinicola, Parasanguinicola) or delicate and appearing irregular or discordant (Pseudosanguinicola) distributing in a single column (Sanguinicola, Parasanguinicola) or distributing as densely compacted transverse rows along lateral body margin (Pseudosanguinicola). Rosethorn-shaped spines absent. Sensory papillae present (Sanguinicola, Nomasanguinicola, Pseudosanguinicola) or absent (Cladocaecum, Kritsky, Plehniella) bearing cilia (Sanguinicola, Nomasanguinicola, Parasanguinicola, Pseudosanguinicola) or lacking cilia (Cladocaecum, Kritsky, Plehniella), not distributing in regular dorsolateral bands. Anterior sucker a proboscis accommodating the mouth, not separated from body by peduncle, having denticles in 2 columns flanking mouth (Nomasanguinicola) or lacking denticles (Cladocaecum, Kritsky, Parasanguinicola, Plehniella, Pseudosanguinicola, Sanguinicola), having concentric spine rows in small adults (Kritsky, Plehniella) or aspinose (Cladocaecum, Sanguinicola, Parasanguinicola, Pseudosanguinicola); mouth medioventral, subterminal, comprising a minute pore-like opening, with associated toothlike mouth apparatus of Ejsmont (1926) (Pseudosanguinicola, Sanguinicola). Nervous system well developed; dorso- and ventrolateral nerve cords extending for nearly entire body length, meeting to form anterior and posterior nerve commissures; secondary lateral branches obvious (Sanguinicola) or indistinct (Cladocaecum, Kritsky, Nomasanguinicola, Parasanguinicola, Plehniella, Pseudosanguinicola). Pharynx present (Cladocaecum, Kritsky, Nomasanguinicola, Parasanguinicola, Plehniella, Sanguinicola) or absent (Pseudosanguinicola), not highly muscular, not intensely basophilic. Esophagus with anterior and posterior esophageal swellings. Intestine comprising 4 (exceptionally 5) ceca forming an X-shaped configuration (Parasanguinicola, Pseudosanguinicola, Sanguinicola) or having 5 or 6 radial ceca (Kritsky, Nomasanguinicola, Plehniella) or exceptionally having paired anterior ceca plus a medial cecum with numerous branches extending laterally (Cladocaecum), restricted to anterior half of body (Kritsky, Nomasanguinicola, Parasanguinicola, Pseudosanguinicola, Plehniella, Sanguinicola) or extending in posterior half (Cladocaecum), bifurcating at midline, blind ending, not overlapping another cecum.

Testis single, anterior to ovary and genitalia, diffuse, having laterally directed lobes or exceptionally (purportedly) not having lateral-directed lobes (Parasanguinicola), post-cecal (Kritsky, Nomasanguinicola, Parasanguinicola, Pseudosanguinicola, Plehniella, Sanguinicola) or dorsal to cecum (Cladocaecum), in middle one-third of body (Kritsky, Nomasanguinicola, Parasanguinicola, Plehniella, Pseudosanguinicola, Sanguinicola) or posterior half of body (Cladocaecum). Vasa efferentia gathering secondary ducts emanating from lateral margins of testicular lobes, coalescing ventrally along midline, forming large seminal column (Pseudosanguinicola, Sanguinicola) or not forming large seminal column (Cladocaecum, Kritsky, Nomasanguinicola, Parasanguinicola, Plehniella), narrowing medially before connecting to vas deferens; vas deferens straight (Cladocaecum, Kritsky, Nomasanguinicola, Parasanguinicola, Pseudosanguinicola, Sanguinicola) or looping (Plehniella), meeting with cirrus sac. Cirrus sac sinistral, post-ovarian thin walled; internal seminal vesicle present; cirrus everting dorsally in sinistral body half (Plehniella) or everting dorsally near midline (Kritsky, Sanguinicola) or indistinct (Cladocaecum, Nomasanguinicola, Parasanguinicola, Pseudosanguinicola), post-cecal body protuberance associated with cirrus present (Plehniella, Sanguinicola) or absent (Cladocaecum, Kritsky, Nomasanguinicola, Parasanguinicola, Pseudosanguinicola), directing dorsally, medial. Common genital atrium and common genital pore present (Plehniella) or absent (Cladocaecum, Kritsky, Nomasanguinicola, Parasanguinicola, Pseudosanguinicola, Sanguinicola); separate genital pores dorsal, post-cecal female genital pore medial (Cladocaecum, Kritsky, Nomasanguinicola, Pseudosanguinicola, Sanguinicola) or exceptionally (purportedly) sinistral (Parasanguinicola), post-ovarian (Cladocaecum, Kritsky, Nomasanguinicola, Pseudosanguinicola, Sanguinicola) or exceptionally (purportedly) pre-ovarian (Parasanguinicola), aspinose. Auxiliary external seminal vesicle absent.

Ovary medial, post-cecal post-testicular can appear butterfly shaped, having deep lobes (Kritsky, Parasanguinicola, Plehniella) or superficial lobes (Cladocaecum, Nomasanguinicola, Pseudosanguinicola, Sanguinicola), as wide as testicular field (Cladocaecum, Kritsky, Pseudosanguinicola, Plehniella, Sanguinicola) or wider than testicular field (Nomasanguinicola, Parasanguinicola). Oviduct a thin-walled duct emanating from ovary, mediodextral, curving slightly sinistrally; oviducal seminal receptacle present. Vitellarium diffuse, symmetrical, follicular, mostly anterior to genital pores; primary vitelline duct single/unpaired, extending posteriad along midline, not forming a transverse vitelline duct or reservoir, connecting with oviduct in posterior body end proximal to and near oötype. Laurer’s canal absent. Oötype spheroid (Cladocaecum, Kritsky, Pseudosanguinicola, Sanguinicola) or oblong (Nomasanguinicola, Parasanguinicola, Plehniella). Uterus short, having ascending portion only (Parasanguinicola, Pseudosanguinicola, Sanguinicola) or ascending and descending portions (Cladocaecum, Kritsky, Nomasanguinicola, Plehniella), not convoluted, not containing sperm; uterine seminal receptacle absent; uterine eggs thin-shelled, not operculate, triangular or ovoid (Parasanguinicola, Sanguinicola) or spheroid (Cladocaecum, Kritsky, Nomasanguinicola) or indistinct (Plehniella, Pseudosanguinicola), lacking discernible miracidium; metraterm robust and strongly muscular (Plehniella) or indistinct (Cladocaecum, Kritsky, Nomasanguinicola, Parasanguinicola, Pseudosanguinicola, Sanguinicola).

Excretory vesicle Y-shaped or a small gland (Parasanguinicola, Sanguinicola) or indistinct (Cladocaecum, Kritsky, Nomasanguinicola, Plehniella); excretory pore terminal. Maturing in blood or body cavity of later-branching ray-finned freshwater fishes (Teleostei).

Differential diagnosis:

Anterior sucker a proboscis with mouth, not pedunculate, having denticles or lacking denticles or having concentric spine rows in small adults or aspinose; mouth comprising a minute pore-like opening; pharynx present or absent; medial esophageal swelling (pouch) present. Intestine comprising short radial ceca of approximate equal length or having anterior ceca plus a single medial and dendritic posterior cecum. Testis with appendix-like lateral lobes, non-sinusoidal. Genital pores common or separate. Laurer’s canal absent. Vitellarium symmetrical. Maturing in blood and body cavity of later-branching freshwater ray-finned fishes (Teleostei).

Remarks

Sanguinicolids primarily infect later-branching freshwater ray-finned fishes (Teleostei) and differ from all other blood flukes by the combination of having a diminutive anterior sucker, a medial esophageal swelling (pouch), short, radial ceca of approximate equal length or having short anterior ceca plus an elongate, dendritic posterior cecum, testis with appendix-like lateral lobes, and no Laurer’s canal. Warren et al. (2023) treated the taxonomic problems and systematics of Sanguinicola.

Sanguinicola maritimus Nolan and Cribb (2005) is the only sanguinicolid that infects a marine fish. The identity of the molluscan intermediate host is indeterminate but could comprise a littoral euryhaline snail, for example. It has some unique morphological features highlighted by Nolan and Cribb (2005) that could warrant proposal of a new sanguinicolid genus. No nucleotide sequence for this species exists in GenBank.

The original description of Parasanguinicola reported that its type species lacked lateral lobes of the testis and had a pre-ovarian female genital pore lateral to the testis, but these features are dubious and need to be confirmed (Herbert and Shaharom-Harrison, 1995).

Aporocotylidae Odhner, 1912, emended

Accepted genera:

Aporocotyle Odhner, 1900; Deontacylix Linton, 1910; Psettarium Goto and Ozaki, 1929; Paradeontacylix Mcintosh, 1934; Cardicola Short, 1953; Psettaroides Lebedev and Mamaev, 1968; Neoparacardicola Yamaguti, 1970; Metaplehniella Lebedev and Parukhin, 1972; Plethorchis Martin, 1975; Pearsonellum Overstreet and Køie, 1978; Pseudocardicola Parukhin, 1985; Cruoricola Herbert, Shaharom-Harrison, and Overstreet, 1994; Elaphrobates Bullard and Overstreet, 2003; Ankistromeces Nolan and Cribb, 2004; Adelomyllos Nolan and Cribb, 2004; Chaulioleptos Nolan and Cribb, 2005; Phthinomita Nolan and Cribb, 2006; Braya Nolan and Cribb, 2006; Littorellicola Bullard, 2010; Skoulekia Alama-Bermejo, Montero, Raga, and Holzer, 2011; Rhaphidotrema Yong and Cribb, 2011; Primisanguis Bullard, 2012; Cardallagium Yong, Cutmore, Jones, Gauthier, and Cribb, 2018; Allocardicola Yong, Cribb, and Cutmore, 2021; Holocentricola Cutmore and Cribb, 2021.

Diagnosis:

Hermaphroditic, asexual reproduction in estuarine and marine polychaetes, lacking encysted metacercaria or second intermediate host, having migratory schistomulum infecting lymphatic and blood vascular system, having adult infecting blood and body cavity of later-branching ray-finned fishes (Euteleosti). Sporocyst present (Cardicola) or absent (Aporocotyle), fusiform (Cardicola), having few or many cercariae (Cardicola). Redia present (Aporocotyle) or absent (Cardicola); fusiform, having many cercariae. Cercaria non-acetabulate apharyngeate, non-ocellate having spinous anterior sucker, lacking body fin and tegumental body spines discernable gonads or genitalia, tail furcae present and symmetrical with fins (Aporocotyle) or absent (Cardicola); undergoing considerable larval development in vertebrate definitive host. Extrauterine eggs embedding in fish gill epithelium and undergoing considerable larval development; miracidium can hatch and emerge from eggs embedded in fish gill epithelium.

Body of adult strongly dorsoventrally flat (Adelomyllos, Allocardicola, Aporocotyle, Braya, Cardallagium, Cardicola, Chauliolaptos, Crouricola, Deontacylix, Elaphrobates, Holocentricola, Littorellicola, Metaplehniella, Neoparacardicola, Paradeontacylix, Pearsonellum, Plethorchis, Primisanguis, Psettarium, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia) or exceptionally cylindrical/threadlike (Ankistromeces, Phthinomita), spinous; head collar, copulatory bursa, acetabulum, and median esophageal pouch or plicate organ lacking. Tegumental body spines distributing in ventrolateral transverse rows or exceptionally having spine bosses (Aporocotyle only), having distally recurved tip (Cardicola, Elaphrobates, Holocentricola, Littorellicola, Paradeontacylix, Pearsonellum, Skoulekia) or not having distally recurved tip (Adelomyllos, Allocardicola, Ankistromeces, Aporocotyle, Cardallagium, Deontacylix, Phthinomita, Psettarium, Rhaphidotrema); rosethorn-shaped spines present in Paradeontacylix only. Anterior sucker diminutive, spheroid, lacking peduncle, having rows of concentric spines anterior to mouth (Adelomyllos, Allocardicola, Ankistromeces, Aporocotyle, Cardicola, Elaphrobates, Holocentricola, Pearsonellum, Phthinomita, Psettarium) or lacking spheroid anterior sucker (Cardallagium, Littorellicola, Metaplehniella, Neoparacardicola, Paradeontacylix, Psettaroides, Rhaphidotrema, Skoulekia); mouth subterminal, comprising a minute pore-like opening. Nervous system well developed or indistinct (Ankistromeces, Phthinomita); dorso- and ventrolateral nerve cords extending for nearly entire body length, forming anterior and posterior nerve commissures; secondary lateral branches indistinct. Sensory papillae present (Cardallagium, Cardicola, Littorellicola, Psettarium, Primisanguis) or absent (Adelomyllos, Ankistromeces, Aporocotyle, Braya, Chaulioleptos, Cruoricola, Deontacylix, Elaphrobates, Holocentricola, Metaplehniella, Neoparacardicola, Paradeontacylix, Pearsonellum, Phthinomita, Plethorchis, Primisanguis, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia). Pharynx absent. Esophagus enveloped by glandular region (Adelomyllos, Aporocotyle, Braya, Cardallagium, Cardicola, Chaulioleptos, Cruoricola, Elaphrobates, Holocentricola, Littorellicola, Metaplehniella, Neoparacardicola, Paradeontacylix, Pearsonellum, Plethorchis, Primisanguis, Psettarium, Psettaroides, Pseudocardicola, Skoulekia) or not (Ankistromeces, Deontacylix, Phthinomita, Rhaphidotrema); posterior esophageal swelling absent; having a nearly indistinct posterior swelling in (Adelomyllos only).

Intestine comprising 4 ceca (1 pair of anterior ceca plus 1 pair of posterior ceca) forming an X-shaped configuration (Neoparacardicola) or an H-shaped configuration (Adelomyllos, Ankistromeces, Aporocotyle, Braya, Cardallagium, Cardicola, Chaulioleptos, Crouricola, Deontacylix, Elaphrobates, Holocentricola, Littorellicola, Metaplehniella, Neoparacardicola, Paradeontacylix, Pearsonellum, Phthinomita, Plethorchis, Primisanguis, Psettarium, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia) or anterior ceca secondarily lost (Allocardicola), bifurcating at midline, blind ending; posterior ceca not overlapping corresponding cecum, terminating in the posterior one-third of the body (Ankistromeces, Aporocotyle, Braya, Cardicola, Chaulioleptos, Cruoricola, Elaphrobates, Littorellicola, Paradeontacylix, Primisanguis, Psettarium, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia) or terminating in the middle one-third of the body (Adelomyllos, Allocardicola, Cardallagium, Deontacylix, Holocentricola, Metaplehniella, Pearsonellum, Phthinomita, Plethorchis) or terminating in the anterior one-third of the body (Neoparacardicola). Genitalia restricted to posterior one-fourth of body, post-cecal (Adelomyllos, Allocardicola, Ankistromeces, Braya, Cardicola, Chaulioleptos, Cruoricola, Deontacylix, Elaphorbates, Holocentricola, Metaplehniella, Neoparacardicola, Phthinomita, Plethorchis, Primisanguis, Psettarium, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia) or inter-cecal (Aporocotyle), post-testicular (Ankistromeces, Allocardicola, Aporocotyle, Braya, Cardicola, Cruoricola, Deontacylix, Elaphorbates, Holocentricola, Metaplethniella, Pearsonellum, Plethorchis, Primisanguis, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia) or inter-testicular (Adelomyllos, Chaulioleptos, Neoparacardicola, Phthinomita, Psettarium), anterior to oötype (Adelomyllos, Allocardicola, Ankistromeces, Aporocotyle, Cardallagium, Chaulioleptos, Cruoricola, Littorellicola, Neoparacardicola, Paradeontcylix, Pearsonellum, Phthinomita, Plethorchis, Primisanguis, Psettarium, Pseudocardicola, Rhaphidotrema, Skoulekia) or posterior to oötype (Cardicola, Elaphrobates, Braya, Deontacylix, Metaplehniella) or female genitalia is anterior to oötype and male genitalia is posterior to oötype (Holocentricola, Psettaroides).

Testis(es) single (Allocardicola, Ankistromeces, Braya, Cardallagium, Cardicola, Cruoricola Elaphrobates, Holocentricola, Metaplehniella, Pearsonellum, Primisanguis, Psettaroides, Skoulekia) or multiple (Adelomyllos, Aporocotyle, Chaulioleptos, Deontacylix, Littorellicola, Neoparacardicola, Paradeontacylix, Phthinomita, Plethorchis, Psettarium, Pseudocardicola, Rhaphidotrema), dendritic (Metaplehniella, Neoparacardicola) or rectangular (Adelomyllos, Allocardicola, Ankistromeces, Braya, Cardallagium, Cardicola, Chaulioleptos, Crouricola, Deontacylix, Elaphrobates, Holocentricola, Pearsonellum, Phthinomita, Primisanguis, Psettarium, Psettaroides, Rhaphidotrema, Skoulekia) or ovoid (Pseudocardicola) or cobblestonelike (Aporocotyle, Paradeontacylix, Plethorchis, Littorellicola); anterior testis(es) pre-ovarian occupying space anterior to genitalia, inter-cecal (Ankistromeces, Aporocotyle, Braya, Cardicola, Chaulioleptos, Cruoricola, Elaphorbates, Littorellicola, Paradeontacylix, Pearsonellum, Phthinomita, Primisanguis, Psettarium, Psettaroides, Pseudocardicola, Skoulekia) or post-cecal (Adelomyllos, Allocardicola, Cardallagium, Deontacylix, Holocentricola, Metaplehniella, Neoparacardicola, Plethorchis, Rhaphidotrema); posterior testis post-ovarian occupying space posterior to genitalia, post-cecal. Vasa efferentia coalescing in posteroventral region of testis, uniting into a vas deferens in anterior or posterior margin of testis; vas deferens straight to sinuous. Auxiliary external seminal vesicle present only in Pearsonellum only. Cirrus sac sinistral, pre-ovarian (Aporocotyle, Neoparacardicola), post-ovarian (Adelomyllos, Allocardicola, Ankistromeces, Braya, Cardallagium, Cardicola, Chaulioleptos, Cruoricola, Deontacylix, Elaphrobates, Holocentricola, Littorellicola, Metaplehniella, Paradeontacylix, Pearsonellum, Phthinomita, Plethorchis, Primisanguis, Psettarium, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia), thin-walled (the cirrus sac can be indistinct without differential interference contrast optical components); internal seminal vesicle present; cirrus appearing appendix-like when everted; posterolateral body protuberance associated with cirrus present (Ankistromeces, Cardallagium, Cardicola, Holocentricola, Littorellicola, Metaplehniella, Neoparacardicola, Primisanguis, Psettarium, Psettaroides, Skoulekia) or absent (Adelomyllos, Allocardicola, Aporocptyle, Braya, Chaulioleptos, Cruoricola, Deontacylix, Elaphrobates, Paradeontacylix, Pearsonellum, Plethorchis, Pseudocardicola, Rhaphidotrema), directing dorsally. Common genital atrium absent. Genital pores separate, dorsal, sinistral, aspinose, or exceptionally having stylet with Rhaphidotrema only.

Ovary medial (Adelomyllos, Allocardicola, Ankistromeces, Cardallagium, Cardicola, Chaulioleptos, Cruoricola, Deontacylix, Elaphrobates, Littorellicola, Metaplehniella, Paradeontacylix, Pearsonellum, Phthinomita, Plethorchis, Psettarium, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia) or lateral to midline (Braya, Neoparacardicola, Primisanguis), post-testicular (Ankistromeces, Cardallagium, Cardicola, Cruoricola, Deontacylix, Elaphrobates, Littorellicola, Metaplehniella, Paradeontacylix, Pearsonellum, Phthinomita, Plethorchis, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia) or inter-testicular (Adelomyllos, Chaulioleptos, Neoparacardicola, Phthinomita, Psettarium), having smooth margins (Allocardicola, Ankistromeces, Aporocotyle, Littorellicola, Pearsonellum, Phthinomita, Rhaphidotrema, Skoulekia) or having lobed margins (Adelomyllos, Braya, Cardalllagium, Cardicola, Chaulioleptos, Cruoricola, Deontacylix, Elaphorbates, Holocentricola, Neoparacardicola, Paradeontacylix, Plethorchis, Primisanguis, Psettarium, Psettaroides, Pseudocardicola) or purportedly dendritic (Metaplehniella), having well-differentiated refractive acini. Oviduct a thin-walled duct emanating from ovary, medial (Aporocotyle, Metaplehniella) or dextral (Adelomyllos, Allocardicola, Ankistromeces, Braya, Cardallagium, Cardicola, Chaulioleptos, Cruoricola, Deontacylix, Elaphrobates, Holocentricola, Littorellicola, Neoparacardicola, Paradeontacylix, Pearsonellum, Phthinomita, Plethorchis, Primisanguis, Psettarium, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia), straight to sinuous or exceptionally looping in Pseudocardicola only; oviducal seminal receptacle present (Braya, Cardicola, Deontacylix, Elaphrobates, Littorellicola, Paradeontacylix, Pearsonellum) or absent (Adelomyllos, Allocardicola, Ankistromeces, Aporocotyle, Cardallagium, Chaulioleptos, Cruoricola, Holocentricola, Metaplehniella, Neoparacardicola, Phthinomita, Plethorchis, Primisanguis, Psettarium, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia). Vitellarium diffuse or densely compacted, extending from cecal bifurcation to genitalia, symmetrical (Allocardicola, Aporocotyle, Braya, Cardallagium, Cardicola, Cruoricola, Deontacylix, Holocentricola, Littorellicola, Metaplehniella, Neoparacardicola, Paradeontacylix, Primisanguis, Psettarium, Psettaroides, Pseudocardicola, Rhaphidotrema) or asymmetrical (Adelomyllos, Ankistromeces, Aporocotyle, Chauliolaptos, Elaphrobates, Pearsonellum, Phthinomita, Plethorchis, Skoulekia), mostly anterior to genital pores, coalescing along midline (Adelomyllos, Allocardicola, Braya, Cardallagium, Cardicola, Cruoricola, Deontacylix, Elaphrobates, Holocentricola, Littorellicola, Metaplehniella, Neoparacardicola, Paradeontacylix, Primisanguis, Psettarium, Psettaroides, Rhaphidotrema, Skoulekia) or coalescing laterally (Ankistromeces, Aporocotyle, Chaulioleptos, Pearsonellum, Phthinomita, Plethorchis, Pseudocardicola); primary vitelline duct single/unpaired, extending posteriad along midline (Adelomyllos, Braya, Cardicola, Cruoricola, Elaphrobates, Metaplehniella, Neoparacardicola, Psettaroides, Skoulekia) or dextral (Ankistromeces, Aporocotyle, Cardallagium, Chaulioleptos, Deontacylix, Holocentricola, Littorellicola, Paradeontacylix, Pearsonellum, Phthinomita, Plethorchis, Primisanguis, Psettarium, Pseudocardicola, Rhaphidotrema), not forming a transverse vitelline duct or reservoir, uniting with oviduct in posterior body end near oötype. Laurer’s canal absent. Oötype spheroid to oblong. Uterus elongate and convoluted (Adelomyllos, Allocardicola, Aporocotyle, Braya, Cardallagium, Cardicola, Cruoricola, Deontacylix, Elaphrobates, Holocentricola, Littorellicola, Metaplehniella, Paradeontacylix, Pearsonellum, Primisanguis, Psettarium, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia) or straight (Ankistromeces, Chaulioleptos, Phthinomita, Plethorchis), initially extending anteriad from oötype (Adelomyllos, Allocardicola, Ankistromeces, Aporocotyle, Cardallagium, Cruoricola, Deontacylix, Holocentricola, Littorellicola, Metaplehniella, Pearsonellum, Phthinomita, Psettarium, Psettaroides, Pseudocardicola, Rhaphidotrema, Skoulekia) or initially extending slightly posteriad (Braya, Cardicola, Elaphrobates, Paradeontacylix, Primisanguis), curving anteriad along midline, curving posteriad at level of ovary (Adelomyllos, Allocardicola, Braya, Cardallagium, Cardicola, Chaulioleptos, Cruoricola, Elaphrobates, Holocentricola, Littorellicola, Metaplehniella, Paradeontacylix, Phthinomita, Primisanguis, Psettarium, Psettaroides, Rhaphidotrema, Skoulekia), curving posteriad at level of testis (Ankistromeces, Aporocotyle, Deontacylix, Neoparacardicola, Pearsonellum, Plethorchis, Pseudocardicola) or having ascending and descending portions; uterine seminal receptacle present in Primisanguis only; uterine eggs thin shelled, operculate (Pearsonellum) or non-operculate pliable, elongate (Adelomyllos, Braya, Neoparacardicola, Skoulekia) or spheroid (Ankistromeces, Aporocotyle, Cardallagium, Cardicola, Deontacylix, Elaphorbates, Littorellicola, Pearsonellum, Phthinomita, Primisanguis, Psettarium, Rhaphidotrema) or elliptical to ovoid (Chaulioleptos, Cruoricola, Holocentricola, Paradeontacylix, Plethorchis), lacking discernible miracidium; metraterm weakly muscular.

Excretory vesicle V shaped (Plethorchis) or Y shaped (Adelomyllos, Aporocotyle, Cruoricola, Elaphrobates, Littorellicola, Primisanguis, Psettarium, Rhaphidotrema) or a small gland (Ankistromeces, Braya, Cardicola, Chaulioleptos, Holocentricola, Pearsonellum, Phthinomita); excretory pore terminal. Maturing in blood of principally late-branching ray-finned marine fishes (Euteleostei).

Differential diagnosis:

Anterior sucker diminutive, not bowl-shaped, lacking peduncle, spheroid and having rows of concentric spines anterior to mouth or lacking spheroid anterior sucker; mouth comprising a minute pore-like opening; medial esophageal swelling (pouch) absent; pharynx absent. Lateral tegumental spines distributing in ventrolateral transverse rows or spine bosses (Aporocotyle only), not C shaped. Testis(es) non-sinusoidal lacking appendix-like lateral lobes. Intestine comprising 4 ceca (1 pair of anterior ceca plus 1 pair of posterior ceca) forming an X- or H-shaped configuration; anterior ceca long or secondarily lost (Allocardicola only); posterior ceca comprising a pair of elongate ceca each extending posteriad in parallel with respective body margin and terminating in middle or posterior one-third of body or anterior one-third (Neoparacardicola only). Genital pores sinistral or having sinistral common genital pore. Vitellarium asymmetrical or symmetrical. Laurer’s canal absent. Maturing in blood of principally later-branching ray-finned marine fishes (Teleostei).

Remarks

Aporocotylids primarily infect later-branching marine and estuarine ray-finned fishes (Teleostei) and differ from all other blood flukes by the combination of having a spheroid anterior sucker with concentric rows of circumferential spines or the spheroid anterior sucker is lost in the adult or adults have a diminutive anterior sucker, a sinuous esophagus lacking a medial swelling, an X- or H-shaped intestine having 4 ceca, long anterior ceca, long smooth posterior ceca that extend posteriad in parallel with respective body margin and terminate near the posterior body end, testis(es) that lack appendix-like lateral lobes, no Laurer’s canal, and a sinistral common genital pore or separate genital pores that are sinistral.

Three genera (Chanicola Yong, Cribb, and Cutmore, 2021; Spirocaecum Yong, Cribb, and Cutmore, 2021; Balistidicola Cutmore and Cribb, 2022) were recently proposed based on the identities of their definitive hosts, a combination of morphological features common to many genera of Aporocotylidae (sensu stricto, as diagnosed herein), and 28S and ITS2 phylogenetic analyses (Yong et al., 2021; Cutmore and Cribb, 2022). Cutmore and Cribb (2022) asserted that Balistidicola was unique by having short anterior ceca, long posterior ceca, a lanceolate body, a single testis, a post-ovarian uterus, and an oötype that is dextral and anterior to the cirrus sac. The type species of Cardicola (Cardicola cardiocola [Manter, 1947] Short, 1953) has each of those features (Manter, 1947; Short, 1953; Warren et al., 2021). The narrative description as well as Figure 4 of Warren et al. (2021), wherein the type species of Cardicola is redescribed based on the Manter’s holotype and newly collected specimens, clearly shows that this species has each of those features—in particular the oötype is clearly anterior and dextral to the cirrus sac. Further, Cardicola abu Yong, Cutmore, and Cribb, 2018; Cardicola ambrosioi Braicovich, Etchegoin, Timi, and Sardella, 2006; Cardicola coridodacis Manter, 1954; Cardicola currani Bullard and Overstreet, 2004; Cardicola forsteri Cribb, Daintith, and Munday, 2000; Cardicola nonamo Bullard, 2010; Cardicola opisthorchis Ogawa, Ishimaru, Shirakashi, Takami, and Grabner, 2011; Cardicola orientalis Ogawa, Tanaka, Sugihara, and Takami, 2010; Cardicola ahi Yamaguti, 1970 incertae sedis (see Warren et al., 2021); and Cardicola kurochkini (Parukhin, 1976) Bullard and Overstreet, 2006 incertae sedis (see Warren et al., 2021) have this combination of features. Because of these facts, we reassign the type species of Balistidicola to Cardicola as Cardicola corneri (Cutmore and Cribb, 2022) n. comb. as well as reassign Balistidicola yuelao Yong, Cutmore, and Cribb, 2018 as Cardicola yuelao Yong, Cutmore, and Cribb, 2018. The reassignment of the type species of Balistidicola to Cardicola makes the genus a junior subjective synonym of Cardicola.

Regarding Chanicola and Spirocaecum, Yong et al. (2021) stated that these genera were diagnosed with morphological features that do not differentiate them from Cardicola, that is, the diagnoses are not diagnostic. To justify this approach, Yong et al. (2021, p. 1) advocated that, “molecular phylogenetics has shown that emphasizing phenetics alone is unreliable.” These authors detailed their experiential concept of what they called “wastebasket taxonomy” wherein the morphological features they chose failed them in predicting the evolutionary relationships they recovered by their nucleotide-based analyses. We have the antithetical perspective based on available evidence. Our experience is that nucleotide-based phylogenetic reconstruction recovers morphologically similar and diagnosable blood fluke taxa as sharing a common ancestor; correspondingly, the morphological features used by us previously, and herein diagnose and differentiate the monophyletic families (Key) and their genera. Herein, for the lack of functional diagnoses (see previous discussion), we reassign the type species of Chanicola and the type species of Spirocaecum to Cardicola as Cardicola jiigurru Yong, Cutmore, Miller, Wee, and Cribb, 2016 and Cardicola coeptus Nolan and Cribb, 2006, respectively. These reassignments make Chanicola and Spirocaecum junior synonyms of Cardicola. As a result, the additional species are reassigned to Cardicola: Cardicola suni Yong, Cutmore, Miller, Wee, and Cribb, 2016 (formerly Chanicola) as well as the species formerly assigned to Spirocaecum (Cardicola bartolii Nolan and Cribb, 2006; Cardicola covacinae Nolan and Cribb, 2006; Cardicola lafii Nolan and Cribb, 2006; Cardicola tantabiddii Nolan and Cribb, 2006; Cardicola mogilae Brooks, Cribb, Yong and Cutmore, 2017).

We continue to view Cardicola spp. and the Cardicola-like aporocotylids as recently evolved and coradiating with euteleost fishes (Bullard, 2013; Orélis-Ribeiro et al., 2014). This could explain the slight morphological differences between them. Some fish blood fluke genera include species that are morphologically similar but have large nucleotide differences between them (Bullard, 2014; Orélis-Ribeiro et al., 2017; Yong et al., 2018; Warren et al., 2021; Cutmore et al., 2022), and, alternatively, some fish blood flukes that are morphologically easily differentiated have few nucleotide differences (Bullard and Overstreet, 2003; Warren et al., 2021; Yong et al., 2021; Cutmore et al., 2022; Cutmore and Cribb, 2022). Given that there is no direct empirical correlation between morphological similarity/dissimilarity and the corresponding degree of nucleotide similarity for specific lineages of trematodes, we think that it is perilous to assume that tree branch length justifies new genera as suggested by Yong et al. (2021). Without a morphological differential diagnosis, the limits of a genus would be arbitrary (because the tree topology changes upon adding a sequence) and depend on ingroup taxon selection, the marker being used, the methodological details of the phylogenetic analysis, and the outgroup selection.

Key to the families of fish blood flukes based on the morphological diagnoses given previously

• 1a.Having C-shaped lateral tegumental spines each mounted on a muscular peduncle and having non-sinusoidal testis or lacking lateral tegumental spines and having sinusoidal testis; vitellarium symmetrical or asymmetrical, having a dextral primary vitelline collecting duct. Maturing in blood of sharks, rays, and chimaeras (Chondrichthyes) Chimaerohemecidae Yamaguti, 1971
• 1b.Lacking C-shaped lateral tegumental spines; lacking sinusoidal testis; vitellarium asymmetrical or symmetrical 2
• 2a.Anterior sucker robust, bowl shaped, pedunculate; mouth comprising a large opening; pharynx present or absent; lateral tegumental spines spike-like (not C shaped, lacking recurved tip; can be lost in adults of some species); Laurer’s canal present or absent; common genital atrium and pore present 3
• 2b.Anterior sucker neither bowl shaped nor pedunculate; mouth comprising a minute pore-like opening; Laurer’s canal absent; genital pores separate or common 4
• 3a.Anterior sucker having minute spines on inner anteroventral surface in adults; tegumental spines distributing in ventrolateral transverse rows, with distal end having a sharp tip protruding slightly from tegument; pharynx present; intestine inverse U shaped (anterior ceca absent) with posterior ceca terminating near the excretory bladder; 6 testes (5 pre-ovarian testes plus 1 post-ovarian testis), inter-cecal having shallow lobes; common genital pore dextral, post-ovarian. Maturing in blood of sturgeons and paddlefishes (Acipenseriformes) Acipensericolidae n. fam.
• 3b.Anterior sucker aspinose in adults; tegumental spines not distributing in ventrolateral transverse rows, with distal end having a sharp tips enveloped by tegument (adults of Elopicola spp. are aspinose); pharynx present or absent; intestine generally inverse U shaped; anterior ceca short or nearly indistinct; posterior ceca terminating at level of testis (Elopicola)/anterior-most testis (Paracardicoloides); 2 deeply lobed testes (1 pre-ovarian and inter-cecal testis plus 1 post-ovarian and post-cecal testis) (Paracardicoloides) or 1 inter-cecal deeply lobed testis (Elopicola); common genital pore sinistral, pre-ovarian. Egg in gill epithelium of fish host having elongate polar filaments. Maturing in blood of elopomorphs (Teleostei: Elopomorpha) Elopicolidae n. fam.
• 4a.Medial esophageal swelling (pouch) present; intestine comprising short radial ceca of approximate equal length or having anterior ceca plus a single medial and dendritic posterior cecum; testis with appendix-like lateral lobes; genital pores common or separate. Maturing in blood and body cavity of later-branching freshwater ray-finned fishes (Teleostei) Sanguinicolidae Poche, 1926
• 4b.Medial esophageal swelling (pouch) absent; pharynx absent; intestine X or H shaped; anterior ceca long or secondarily lost (Allocardicola); posterior ceca comprising a pair of elongate ceca each extending posteriad in parallel with respective body margin and terminating in middle or posterior 1/3 of body or in anterior 1/3 of body (Neoparacardicola); testes lacking appendix-like lateral lobes; genital pores sinistral or having sinistral common genital pore. Maturing in blood or body cavity of later-branching marine ray-finned fishes (Teleostei) Aporocotylidae Odhner, 1912

Phylogenetic remarks

Our 28S phylogeny recovered the fish blood flukes as monophyletic and sister to the remaining blood flukes infecting turtles and homeotherms (Fig. 1; Bullard et al., 2019; Bullard and Dutton, 2022). Each of the morphologically diagnosed families was recovered as monophyletic. This tree resembles most previously published blood fluke trees and comprises the largest taxon sampling for fish blood flukes to date. Acipensericolidae was recovered as a sister to all other fish blood flukes, with 2 main clades comprising Chimaerohemecidae + Sanguinicolidae and Elopicolidae + Aporocotylidae.

DISCUSSION

Testing the so-called “deep cophyly” between blood flukes and their craniate hosts remains a key question in the natural history of blood flukes, including that for the schistosomes (Bullard and Dutton, 2022). Because the natural history of the fish blood flukes explains the origin of the schistosomes, their systematics and host–parasite relationships are of broad interest. Smith (1972) and Maillard (1982) asserted that fish blood flukes showed no evidence of “strict phylogenetic host specificity” at the level of genus or species. However, our current understanding of the fish blood flukes is that (1) morphologically similar, phylogenetically related fish blood flukes mature in phylogenetically related fish lineages (see Key), (2) the branching order (based on 28S phylogenetic analyses) for fish blood flukes does not mirror that for craniates (i.e., the chimaerohemecids are not sister to all blood flukes) but does indicate that marine ray-finned fishes are a late-branching lineage among fish blood flukes, and that (3) the phylogenetic position of these major fish blood fluke lineages (each of which is robustly monophyletic) remains unsettled (Fig. 1; Table I). This new understanding derives mainly from discoveries of fish blood flukes infecting chondrichthyans, early-branching actinopterygians, and other freshwater fishes; all of which previously were underexplored for blood fluke infections (Orélis-Ribeiro et al., 2013, 2014, 2017; Orélis-Ribeiro and Bullard, 2015, 2016; Warren et al., 2017, 2019, 2020, 2021, 2023; Warren and Bullard, 2019, 2021). Further, although few fish blood fluke life cycles are known, 11 from freshwater fishes and 8 from marine fishes (Warren and Bullard, 2019; Cutmore et al., 2022; Table II), the fish blood flukes appear to show some level of specificity (and perhaps some level of cophyly) to/with their molluscan first intermediate hosts (Cribb et al., 2017; Warren and Bullard, 2019; Cutmore et al., 2022).

The blood flukes (Schistosomatoidea) seem to be an old lineage because they mature in collectively all major craniate lineages and exploit a large diversity of intermediate hosts. They mature in hosts belonging to the earliest-branching extant jawed vertebrates (Chondrichthyes), to the early-branching actinopterygians (Acipenseriformes), to the late-branching ray-finned fishes (Tetraodontiformes), and freshwater and marine turtles and homeotherms (birds and mammals). Further, they collectively exploit the greatest diversity of invertebrate hosts of any trematode lineage; undergoing asexual reproduction in bivalves and snails but also polychaetes (Table II). Alternatively, but seemingly less likely, is that the blood flukes recently evolved and rapidly colonized and radiated across the craniate tree of life. Extinct early-branching gnathostome lineages could have harbored blood fluke infections, and those blood flukes would comprise key missing links to piece together the natural history of the blood flukes. For example, a blood fluke could have infected any number of extinct fish lineages (pteraspidomorphs, anaspidomorphs, thelodontomorphs, osteostracomorphs, placodermiomorphs) or the 47 extinct lineages (families and orders) of chondrichthyans (Nelson et al., 2016). However, unless the lateral tegumental spines, for example, are discovered in a fossil fish belonging to 1 of these lineages, we could never know. The most alluring craniates that remain to be explored for blood fluke infections are the extant jawless fishes (monophyletic Agnatha) and stem sarcopterygians (lungfishes [Dipnoi] and coelacanths [Coelacanthomorpha]). Despite having examined limited numbers of lampreys and lungfishes (S. A. Bullard, pers. obs.), no infection has been detected so far. Further, the extant agnathans, lungfishes, and coelacanths are each but a relic of a previously more species-rich lineage. Perhaps this theorized extinction of some fish, turtle, and tetrapod blood fluke lineages explains why robust clades of blood flukes are unsettled regarding their phylogenetic position.